Fluorescent proteins are widely used as fusion tags to detect protein expression in vivo. To become fluorescent, these proteins must undergo chromophore maturation, a slow process with a half-time of 5 to >30 min, which causes delays in real-time detection of protein expression. Here, we engineer a genetically-encoded fluorescent biosensor to enable detection of protein expression within seconds in live cells. This sensor for transiently-expressed proteins (STEP) is based on a fully matured but dim green fluorescent protein in which pre-existing fluorescence increases 11-fold in vivo following the specific and rapid binding of a protein tag (Kd 120 nM, kon 1.7 x 10^5 M-1s-1). In live E. coli cells, our STEP biosensor enables detection of protein expression twice as fast as the use of standard fluorescent protein fusions. Our biosensor opens the door to the real-time study of short-timescale processes in research model animals with high spatiotemporal resolution.

中文翻译：

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基因编码的荧光生物传感器可快速检测蛋白质表达

荧光蛋白被广泛用作融合标签，以检测体内蛋白表达。为了变成荧光，这些蛋白质必须经过发色团成熟，这是一个缓慢的过程，半衰期为5到> 30分钟，这导致蛋白质表达的实时检测延迟。在这里，我们设计了一种基因编码的荧光生物传感器，以使能够在数秒内检测到活细胞中的蛋白质表达。这种用于瞬时表达蛋白（STEP）的传感器基于完全成熟但呈暗绿色的荧光蛋白，在该蛋白中，预先存在的荧光在特异性和快速结合蛋白标签后在体内会增加11倍（Kd 120 nM，kon 1.7 x 10 ^ 5 M-1s-1）。在活的大肠杆菌细胞中，我们的STEP生物传感器能够使蛋白质表达的检测速度比使用标准荧光蛋白质融合体快两倍。